| Phytochemicals naturally present in fruits and vegetables that possess both antioxidant and anti-inflammatory properties which have been attributed to many health beneficial effects like preventing cardiovascular diseases(CVD),cancer and other degenerative diseases,as well as delaying aging.Purple carrots and purple potatoes are widely concerned because they are rich in phytochemicals like phenolics and carotenoids.Phenolics and carotenoids are two common phytochemicals with biological activity and co-existed in fruits and vegetables.Phenolics and carotenoids are usually decomposed,metabolized and exerted their effects together in the human digestive system.Whether these phytochemicals interact with each other in the process of digestion,absorption and functioning is still unclear.Based on the characteristics of purple carrots and purple potatoes containing phenolics and carotenoids,our study extracted and analyzed the phenolic and carotenoid components in purple carrots and purple potatoes and compared with normal carrots and potatoes to explore the anti-inflammatory and antioxidant interactions of these components.A co-culture cell model consisted of intestinal epithelial cells(Caco-2)and vascular endothelial cells(EA.hy 926)was established.The transepithelial ability to cross the epithelial cell monolayer,and the anti-inflammatory effects of the phenolic and carotenoid components from purple carrots and purple potatoes were measured in the Caco-2/EA.hy 926 co-culture cell models.The typical anthocyanin and carotenoid standards which could enter to the circulatory system were used to explore the antioxidant interaction and molecular mechanisms in H9c2 cells.And the anti-inflammatory interactions and molecular mechanisms about the final fermentation products of undigested phytochemicals in the large intestine(benzoic acid,phenylacetic acid,and phenylpropionic acid)and short-chain fatty acids(butyric acid)were studied.The major results are concluded as follows:1.Analyzed and identified the phenolic and carotenoid components in purple carrots and compared them with normal carrots.It was found that the total phenolics content(15.49±0.82 mg/g DW)in purple carrots is 8 times higher than that of normal carrots(1.86±0.26 mg/g DW).The total anthocyanins content in purple carrots is 5.52±0.38 mg/g DW,and the highest anthocyanin is cyanidin-3-xylosyl(ferulose glucosyl)galactoside(7.44±0.18 mg)/g DW).Chlorogenic acid is the most abundant phenolic acid in purple and normal carrots,the contents are 8.06±0.30 mg/g DW and 1.32±0.16 mg/g DW,respectively.The total carotenoids content in normal carrots(1.14±0.03 mg/g DW)is 5 times higher than that of purple carrots(0.23±0.01 mg/g DW),and the main carotenoids are lutein,a-carotene and β-carotene.The content of lutein in purple carrots(54.1 ± 1.84μg/g DW)is higher than that of normal carrots(15.32±0.17μg/g DW).Compared with normal carrots,purple carrots have higher phenolic and carotenoid contents and also stronger antioxidant capacities(ORAC,DPPH and FRAP).2.Analyzed and identified the phenolic and carotenoid components in purple potatoes and compared them with normal potatoes.It was found that the total phenolics content(4.75±0.25 mg/g DW)in purple potatoes is 3 times higher than that of normal potatoes(1.60±0.48 mg/g DW).The total anthocyanins content is 4.21±0.34 mg/g DW in purple potatoes,and the highest anthocyanin is petunidin-3-p-coumarylrutinoside-5-glucoside(6.33±0.14 mg/g DW).Chlorogenic acid is the highest content of phenolic acid in potatoes.Purple potato has the highest content of chlorogenic acid dimer(1.62±0.05 mg/g DW),while normal potatoe has the highest content of chlorogenic acid isomer(0.37±0.03 mg/g DW).The total carotenoids content in normal potatoes(3.14±0.02 mg/100g DW)is 5 times higher than that of purple potatoes(0.63±0.01 mg/100g DW).The main carotenoids are neoxanthin,violaxanthin and lutein.The antioxidant capacities(ORAC,DPPH and FRAP)of phenolics components in purple potatoes were stronger than that of normal potatoes,while the antioxidant capacities of carotenoid components in purple and normal potatoes were opposite.3.The phenolics and carotenoids in purple carrots and potatoes showed strong cellular antioxidant capacity and anti-inflammatory effects to inhibit the IL-8 release from Caco-2 cells.And the carotenoid components from purple potatoes are relatively weak in cellular antioxidant capacity and anti-inflammatory capacity in Caco-2 cells.The phenolics and carotenoids from purple carrots mixed with different ratios(phenolics:carotenoids=70:1,30:1,20:1,10:1 and 1:1,w/w)exhibited synergistic antioxidant effects in vitro test,but there is no synergistic effect in the cellular antioxidant and anti-inflammatory assay.The phenolics from purple potatoes mixed with the carotenoid standard(lutein and lycopene,phenolics:carotenoid standard=9:1,7:3,1:1,3:7 and 1:9,w/w)showed a synergistic anti-inflammatory effects when at the ratios of 9:1 and 7:3.The purple potato phenolics mixed with lutein and lycopene showed no synergistic effect of the cellular antioxidant activity.4.The phenolics and carotenoids from purple carrots were able to transport to the basolateral side of the Caco-2 monolayer,and significantly inhibited the secretion of IL-8 and the gene expression of IL-8,VCAM-1 and TNF-α induced by TNF-α in EA.hy 926 cells.Western blot analysis indicated that the phenolics and carotenoids from purple carrots significantly inhibited the phosphorylation of JNK,ERK,p38 and IκB-α in the basal layer EA.hy 926 cells,that mean the phenolics and carotenoids exerte anti-inflammatory functions by blocking NF-κB and MAPK signal pathways.The mixture of phenolics and carotenoids from purple carrots(50 μg/mL+50 μg/mL)had no synergistic anti-inflammatory effects in EA.hy 926 cells.5.The phenolics from purple potatoes and lycopene passed through the Caco-2 monolayer,and synergistically inhibited the secretion of IL-8 and the gene expression of IL-8,VCAM-1 and TNF-α induced by TNF-α in EA.hy 926 cells.The mixture of phenolics from purple potatoes and lycopene(50 μg/mL+2 μg/mL)synergistically inhibits the phosphorylation of JNK,ERK and p38 in EA.hy 926 cells,and showed the synergistic anti-inflammatory effect by inhibiting the NF-κB and MAPK signaling pathways.In addition,their mixture(100 μg/mL:4 μg/mL)has no effect on the transporters GLUT 2 and SR-BI,but synergistically reduced the gene expression of tight junction protein like ZO-1.6.The anthocyanin standard petunidin and the carotenoid standard lycopene synergistically inhibited the oxidative damage induced by H2O2 in H9c2 cells.Petunidin and lycopene were mixed in different ratios(petunidin:lycopene=1:9,3:7,1:1,7:3 and 9:1,w/w).Petunidin and lycopene mixed in the ratio of 9:1(petunidin:lycopene)synergistically inhibited the cellular ROS production and promoted the expression of antioxidant enzymes(SOD,CAT and GSH-Px).Western Blot and RT-PCR experiments revealed that petunidin and lycopene synergistically promoted the phosphorylation of Akt and the expression of NQO1 and HO-1 on the Akt/Nrf2 pathway to achieve an antioxidant synergistic effect.siRNA experiments revealed that petunidn has a greater effect on NQO1,while lycopene on HO-1.7.Petunidin and lycopene were mixed in different ratios(petunidin:lycopene=1:9,3:7,1:1,7:3 and 9:1,w/w).When the ratio is 9:1,petunidin and lycopene synergistically improved ΔΨm,inhibited the release of cytochrome c,promoted the expression of Bcl2 and inhibited the expression of Bax,Caspase-3 and-9,synergistically inhibited H2O2-induced H9c2 cells apoptosis by regulating the mitochondrial apoptosis pathway.8.The end products of phenolics metabolism(benzoic acid,phenylacetic acid and phenylpropionic acid)and butyric acid could inhibit TNF-α-induced IL-8 secretion and gene expression of IL-8,VCAM-1 and TNF-α through MAPK and NF-κB pathways in Caco-2 cells.Butyric acid mixed with the end products of phenolics metabolism(benzoic acid,phenylacetic acid and phenylpropionic acid)in a ratio of 1:1,synergistically inhibited the secretion of IL-8 induced by TNF-α and the gene expression of inflammatory factors such as IL-8,VCAM-1 and TNF-α.And the end products of phenolics metabolism and butyric acid showed the synergistic anti-inflammatory effects by inhibiting the phosphorylation of JNK,ERK,p38 and IKB-α on the MAPK and NF-κB pathways. |
PDF Full Text Request